Many components of the innate immune response mediate the release of inflammatory cytokines during bacterial or viral infection and shock. Our preliminary data support the hypothesis that CD14 and TLR2 participate in the innate immune recognition of whole LCMV, an important potential bioterrorist pathogen. Furthermore, the ability to activate TLR2 may, in fact, be a critical determinant of long-term immunity as this receptor plays an important role in enhancing long-term immunity through the recognition of molecules with adjuvant activity. Substantial evidence suggests that TLR2 initiates immune signaling by forming a receptor complex involving at least one additional TLR. We propose that this combinatorial recognition of a TLR ligand is an important component of the innate immune system, and mediates unique responses by employing heteromeric complexes to distinguish one set of TLR2 ligands from another. In this proposal, we wish to test the hypothesis that TLR2 contributes to the innate immune response to LCMV in conjunction with other TLRs, most likely TLRs1, 6 or possibly 10. We also hypothesize that the precise program of cellular activation following exposure to LCMV is determined by the specific TLR2 co-receptor, which we propose to identify. The scientific goals of this project are divided into three phases. In the first phase, we will test the hypothesis that TLR2 and another TLR (likely 1, 6 or 10) participate in the immune recognition of LCMV. The interactions between TLR2 and its TLR co-receptor will be analyzed by immunological, confocal and immunoelectron microscopy techniques. In a second series of experiments, we will determine if and how the engagement of different combinations of TLR2/TLR receptor complexes results in different programs of cellular activation, by using mouse macrophage microarrays and performing binding assays with LCMV adjuvant proteins discovered in Project 3. In a third, collaborative Aim, we will evaluate whether the endogenous adjuvants of Project 1 engage TLRs and/or whether they work synergistically with proteins from LCMV to enhance the immune response. A better understanding of the interaction between TLR2 and LCMV, the identification of the viral product(s) that mediate this innate response (as outlined in Project 3), and the ability to enhance immune response to LCMV (in collaboration with Project 1) will all be of critical importance for the development of vaccines against LCMV.